P330: TARGETING BTN2A1 BY A UNIQUE ACTIVATING MAB IMPROVES ANTI-TUMOR FUNCTIONS OF VΓ9VΔ2 T CELLS
Authors: A.-C. Le Floch, C. Imbert, A. de Gassart, A. Le Roy, L. Gorvel, N. Vey, A. Anastasio, A. Briantais, N. Boucherit, C. E. Cano, D. Olive
Journal: HemaSphere (2022)

Background: Vγ9Vδ2 T cells are new promising cytotoxic effectors in cancer immunotherapy. In acute myeloid leukemia and in non-Hodgkin lymphomas, Vγ9Vδ2 T cells-based immunotherapy has shown encouraging results both in preclinical models and in early phase clinical trials. But very few data are currently available on susceptibility of acute lymphoblastic leukemia (ALL) cells to Vγ9Vδ2 T cell cytotoxicity. Vγ9Vδ2 T cells are activated by phosphoantigens bound to BTN3A1 on target cells. BTN3A targeting agonist antibody ICT01 is being developed in a multicentric Phase 1 and 2 study called EVICTION by Imcheck Therapeutics. Recently the biology of Vγ9Vδ2 T cells has recently undergone a new paradigm with the identification of BTN2A1 as the direct ligand for Vγ9 chain of γδ TCR. BTN2A1 seems to be mandatory for Vγ9Vδ2 T cell activation but its precise role in modulating functions of Vγ9Vδ2 T cells remains unknown. Aims: Here, we show we show that Vγ9Vδ2 T cells exert cytolytic functions against ALL cell lines and primary ALL blasts and that Vγ9Vδ2 T cell cytotoxic activity is enhanced after treatment with a unique agonist mAb targeting BTN2A1 called 107G3B5. Mechanistically, anti-BTN2A1 enhances interactions between Vγ9Vδ2 T cells and target cells and improves the binding of Vγ9Vδ2 TCR on target cells. Methods: 15 hematological cancer cell lines and PBMC from 17 adults ALL patients at diagnosis (7B-ALL, 7T-ALL and 3Ph+ ALL) were tested in functional assays. We quantified the relative surface expression of BTN2A (using 107G3B5 and 7.48 mAbs) and BTN3A (using 20.1 and 108.5 mAbs) on cell lines and primary ALL blasts. In parallel, allogenic Vγ9Vδ2 T cells functions against cell lines and primary ALL blasts were evaluated. ALL samples were also tested for their expansion capacities after 14 days of PBMC culture in presence of Zoledronate. Interactions of Vγ9Vδ2 T cells with target cells was investigated using a 3D imaging in real-time by holo-tomographic microscopy. Binding assays were realized using a recombinant tetramerized Vγ9Vδ2 TCR. Results: We showed that Vγ9Vδ2 T cells exert spontaneous cytotoxicity against hematological cell lines and primary ALL blasts with a heterogeneous susceptibility depending on the target. We demonstrated that anti-BTN2A1 agonist mAb (107G3B5) significantly enhanced Vγ9Vδ2 T cells mediated apoptosis, in comparison to control condition. This effect was increased over time even for the less spontaneously susceptible cells. We confirmed these observations with autologous Vγ9Vδ2 T cells expanded from 4 ALL patients at diagnosis for which effector functions were increased after treatment with 107G3B5. In live microscopy, 107G3B5 enhanced interactions between target cells cocultured during 24h with Vγ9Vδ2 T cells. Finally, we observed that 107G3B5 strongly increased binding of a recombinant Vγ9Vδ2 TCR to target cells. Summary/Conclusion: Here, we demonstrated that targeting BTN2A1 led to improve Vγ9Vδ2 T cell antitumor response at molecular, cellular and functional levels. Our results highlighted that Vγ9Vδ2 T cells exert cytolytic functions against ALL cells, both in allogenic and autologous setting and demonstrated that BTN2A1 targeting with our unique agonist mAb could potentiate effector activities of Vγ9Vδ2 T cells against ALL blasts. These results indicate that the sensitization of leukemic cells can be induced by activation BTN3A as reported previously as well as with BTN2A1 mAbs. These findings could be of great interest for the design of innovative Vγ9Vδ2 T cells-based immunotherapy strategies for treating ALL that could be extended to other cancer types.